Biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (P(HB-co-HHx)) from butyrate using engineered Ralstonia eutropha

Abstract

Polyhydroxyalkanoates (PHAs), a promising family of bio-based polymers, are considered to be alternatives to traditional petroleum-based plastics. Copolymers like poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (P(HB-co-HHx)) have been shown to exhibit favorable physical and mechanical properties, due to decreased crystallinity resulting from the presence of medium-chain-length 3-hydroxyhexanoate (3HHx) monomers. In this study, we produced P(HB-coHHx) using engineered Ralstonia eutropha strains containing deletions of the acetoacetyl-CoA reductase (phaB) genes and replacing the native PHA synthase with phaC2 from Rhodococcus aetherivorans I24 and by using butyrate, a short-chain organic acid, as the carbon source. Although the wild-type R. eutropha did not produce P(HB-co-HHx) when grown on mixed acids or on butyrate as the sole carbon source, we are able to produce polymer containing up to 40 wt% 3HHx monomer with the aforementioned engineered R. eutropha strains using various concentrations of just butyrate as the sole carbon source. This is the first report for the production of P(HB-co-HHx) copolymer in R. eutropha using butyrate.

Improved artificial pathway for biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) with high C6-monomer composition from fructose in Ralstonia eutropha

“…1 (A)). Very recently, Jeon et al (2014) reported similar effects of deletion of the three PhaB homologs on P(3HB-co-3HHx) biosynthesis from butyrate by recombinant R. eutropha. In the previous results with R. eutropha PHB À 4 expressing phaC from A. caviae and ccr Sc (Fukui et al, 2002), the slight incorporation of 3HHx into PHA on fructose was thought to result from the weak expression of phaB1 caused by the polar effects of the non-sense mutation in phaC on chromosome 1 of this mutant (Mifune et al, 2008).…”

Characterization of poly-3-hydroxybutyrate (PHB) produced from Ralstonia eutropha using an alkali-pretreated biomass feedstock

“…Poly-3-hydroxybutyrate (PHB) is a member of the PHA family and is the best-characterized polymer. The properties of pure PHB such as hydrophobicity, inertness, thermoplastic processability, a relatively high melting point, and optical purity make PHB an attractive alternative to synthetic plastics [2,4]. The materials such as PHB have other advantages over synthetic plastics including biodegradability (to water and carbon dioxide or methane) by a variety of microorganisms under anaerobic conditions in many ecosystems and the possibility of production from renewable sources [5,6].…”

Regulation of 3-hydroxyhexanoate composition in PHBH synthesized by recombinant Cupriavidus necator H16 from plant oil by using butyrate as a co-substrate

“…To increase the 3HH composition, recombinant C. necator strains lacking the phaB genes (phaB1, B2, and B3) have been constructed. Since these mutant strains produce PHBH at low levels, deletion of phaB is counterproductive for industrial-scale PHBH biosynthesis from an economic perspective (18,19). On the other hand, Tsuge et al (20) successfully modified the PHA synthase enzyme (PhaC Ac NSDG) that showed enhanced incorporation of longer 3-hydroxyalkanoate units such as (R)-3HH-CoA into a PHA.…”